Fastx_reverse_complement:Found invalid nucleotide sequence

[Jean-Philippe]

Hello!
I am working on making a pipeline to quickly align and analyse my Sanger sequencing data (16S).
Birelfy, I start from .ab1 files → seperate the reverse and forward → ab1 to FASTQ → seqtk_trimfq → FASTQ Groomer.
I run in a problem when I run Reverse-complement tool. The pipeline works if I use sequences without ambigous nucleotides.
The error message is the following :
If I use sequences with ambigous nucleotides (such as R, W or Y, but excluding N), I get this error message : Fastx_reverse_complement:Found invalid nucleotide sequence.
Is there a way to fix this situation?

Thank you!

[jennaj]

Welcome, @Jean-Philippe

If you have non-specific bases that the FastX tools cannot process, you could convert all of those to Ns. Other tools might present with the same issue, too, so maybe review what those downstream tools are expecting when deciding.

Or, you could try a different tool to process the reverse compliment. I can’t remember how strict Manipulate FASTQ is but it seems worth trying.

Some related resources:

• Getting Data into Galaxy
• Datatype FAQs Galaxy FAQs
• The Quality Control tutorial here has a quick overview with screenshots → Sequence analysis / Tutorial List
• Or choose a tutorial from here → GTN Materials Search (query=16s)
• Galaxy includes a lot of ways to custom manipulate data files. You could convert to a tabular format, make custom changes, then convert back to fastq. → GTN Materials Search (query=olympics)

Hope this gives some ideas!